The substructure of tight junctions is under investigation by direct freezing techniques that avoid any chemical fixation, and serve to increase the resolution of its individual components. The tight junction is made of rod-shaped structures embedded in the central lipophilic domain of each of its paired component membranes. This view revises the earlier view that the tight junction is a row of intramembrane particles in register with a similar row in an adjacent membrane. These results are expected to lead to an understanding of how tight junctions serve in the blood-brain barrier system to prevent small charged solutes from entering the brain.